The purpose of this FIRST award grant application is to determine if endogenous testicular opioids mediate the effects of alcohol on two fundamental aspects of male reproductive function: Testosterone secretion and testicular interstitial fluid (TIF) formation. In preliminary studies in rats, alcohol stimulated testicular B-endorphin secretion and alcohol and opioids suppressed both testicular steroidogenesis and TIF volume. The sites and mechanisms of action of these effects will be examined with special emphasis on the hypothesis that endogenous testicular opioids mediate the in vivo inhibitory effects of alcohol on testicular function. Two specific aims will address these questions. First, the effects of alcohol and opioids on testicular function will be compared and the role of endogenous testicular opioids in these effects will be examined. Acute and chronic systemic effects and intratesticular effects of alcohol, opioids, and opioid antagonists on TIF volumes and the concentrations of testosterone and immunoreactive 3-endorphin (i-BetaE) in TIF will be tested. Similarities between alcohol and opioid effects, cross-tolerance between alcohol and opioids, blockade of alcohol's effects by opioid antagonists, and the effects of alcohol on other endogenous testicular opioids will indicate that endogenous opioids mediate alcohol's testicular effects. The relationships between the effects of alcohol and opioids on TIF volumes, TIF formation, testicular access of nutrients and hormones needed for steroidogenesis and spermatogenesis, and testicular function will be examined by measuring TIF formation, testicular uptake of labeled gonadotropins, testicular oxygen tension, and TIF concentrations of glucose and cholesterol after alcohol and opioid treatments. Second, other possible mechanisms of action of alcohol and its interactions with opioids in testicular steroidogenesis and TIF formation will be examined. These mechanisms, chosen on the basis of the preliminary results and previous literature, include the roles of vasoactive substances such as calcium, nitric oxide, and catecholamines and the roles of corticotropin releasing factor, corticosterone, vasopressin, excitatory amino acids, and testicular luteinizing hormone-releasing hormone in the testicular effects of alcohol and opioids. In vivo systemic and intratesticular injections and in vitro tests of active agents followed by measurements of testosterone secretion and TIF volumes will determine if alcohol and opioids exert similar testicular effects through the same sites and mechanisms of action. The health relatedness of this project will include the definition of a previously undescribed and deleterious effect of alcohol on TIF formation and new insight into testicular endocrinology, paracrinology, and autocrinology important for understanding and treating alcohol- and other drug-induced male infertility. It will also define another important interaction between alcohol and endogenous opioid systems important for understanding the role of opioids in alcohol's effects.